Nov. 5 , 1955
NOTES
riboflavin from the green fermentation product. T h e latter compound was then easily isolated by filtration and was washed with methanol and acetone. The compound (600 riig.) thus obtained remained green in the dry state, but upon exposure to air in the presence of water i t was readily oxidized t o an orange substance which from chromatographi; analysis appears t o be identical with 6,7-dimethyl-9-(2 hydroxyethyl) isoalloxazine. Kuhn and StroheleG reported that one mole of half-reduced flavin could form quinhydrone-like complexes with one mole of flavin (chloroflavin, light green) with one mole of half-reduced flavin (verdoflavin, dark green) or with one mole of dihydroflavin (rhodoflavin, red). In view of the ready air oxidation t o yellow flavin of the green and red substances observed during the fermentation, it appeared possible that these substances are molecular complexes of the kind observed by Kuhn and Strobele. Evidence in support of this hypothesis was obtained by showing that when 100 mg. of the green compound (equivalent t o 175 y 3 1 of presumed complex) was suspended in 2.0 ml. of water and shaken in air for 4 hours in a Warburg respirometer a t 26", 40 p M of oxygen was consumed. No further oxygen uptake occurred even after shaking overnight. T h e observed oxygen uptake corresponds t o 91% of the theoretical uptake expected from a molecular complex composed of one mole of half-reduced and one mole of oxidized flavin. Further evidence in support of the above hypothesis was obtained by showing that when an alkaline solution of the purified bacterial flavin is treated under anaerobic conditions with a solution of sodium hydrosulfite, a dark green precipitate is formed, corresponding t o the verdoflavin reported by Kuhn. Addition of concentrated acid caused formation of a bright red solution (rhodoflavin), which, on dilution with water and gradual admission of air became light green (chloroflavin). It appears very probable, therefore, that the red and green precipitates observed early in the fermentations are in fact quinhydrone-like complexes of partially reduced 6,7-dimethyl-Q-(2'-hydroxyethyl)-isoalloxazine.
a research program which has had to be abandoned, this method was used very satisfactorily in the iodination of tyrosine derivatives. Good yields of monoiodo derivatives were obtained from 3-nitrotyrosine and o-methyltyrosine and of 3,5-diiodotyrosine from tyrosine.
-
(6) R. Kuhn and R. Strobele, B e i . , 70, 753 (1937).
LABORATORY OF CELLULAR PHYSIOLOGY SATIONAL HEARTINSTITUTE KATIONAL INSTITUTES OF HEALTH DEPARTMENT O F HEALTH, EDC'CATION AND BETHESDA 14, MARYLAND
n'ELFARE
The Iodination of Tyrosine and its Derivatives BY L. JURD' RECEIVEDJUNE21, 1955
Early attempts to iodinate tyrosine in various alkaline media were generally unsatisfactory. In recent years, however, tyrosine has been converted into 3,5-diiodotyrosine in good yields by reaction with iodine monochloride2 and with iodine and ethla mine.^ Bauer and Strauss4reported t h a t iodine did not react with 3-nitrotyrosine in alkaline solutions or in the presence of mercuric oxide. With iodine monochloride, they obtained 3-iodo-&nitrotyrosine in small yield. A new method was developed recently for the iodination of phenols and aromatic ether^.^ This method involved the reaction of the aromatic compound with iodine and hydrogen peroxide in the presence of a strong mineral acid. As part of (1) U.S. Department of Agriculture, Pasadena, California. (2) (a) P. Block, Jr., and G. Powell, THISJOURNAL, 65, 1430 (1943); (b) E. T. Borrows, J. C . Clayton and B . A. Hems, J . Chem. Soc., 5185 (1949). (3) J. H. Barnes, E. T. Borrows, J. Folks, B . A. Hems and A. G. Long, i b i d . , 2824 (1950). (4) (a) H. Bauer and E. Strauss, B e y . , 68B,1108 (1935); (b) 69, 245 (1936). ( 5 ) (a) L. Jurd, Azrslrnlion J . Sci. Research, SA,596 (1940); (b) SA, 587 (1950).
5747
Experimental6 3,5-Diiodotyrosine.-Powdered iodine (2.8 9.) was suspended in a solution of tyrosine (2.0 g . ) in glacial acetic acid (14.0 cc.) and 36 N hydrochloric acid (8.0 cc.). Thirty per cent. hydrogen peroxide solution was then added in small portions with shaking during five minutes until the iodine color had disappeared, the temperature of the reaction being maintained a t 60-65". Approximately 1.3 cc. of the hydrogen peroxide solution was required. The yellow solution was cooled and diluted successively with water (10 cc.), 0.880 M ammonia solution (9.0 cc.), and 10% sodium hydrogen sulfite solution ( 5 cc.). 0.880 M ammonia solution was then added dropwise until crystallization began. 3,5Diiodotyrosine separated in flat, almost colorless needles. It was collected, washed with water and alcohol and airdried, m.p. 198" (lit. 201' (cor.)2a)(3.4 g., 71%). 3-Iodo-5-nitrotyrosie.-Concentrated nitric acid (3.0 cc.) was added to a suspension of 3-nitrotyrosine (6.0 9.) and powdered iodine (3.4g.) in 95% alcohol (45 cc.). Thirty per cent. hydrogen peroxide solution was then added in 0.5-cc. portions until the color of iodine had disappeared. 4.0 cc. of hydrogen peroxide solution was required and the heat of the reaction maintained the temperature a t 45-50' throughout the addition. The reaction mixture was heated t o 70" for five minutes, diluted with water (30 cc.) and treated with concentrated ammonia, added dropwise, until a heavy yellow solid separated. After standing a t 0' for two hours, the solid was collected, washed with a small quantity of water and alcohol and heated under reflux with alcohol (30 cc.) for ten minutes. On cooling, the crystalline solid was collected. It was dissolved in hot dilute hydrochloric acid and precipitated with ammonia (5.6 g., 60%, m.p. 220'). For analysis the 3-iodo-5-nitrotyrosine was recrystallized from water and separated in goldenyellow needles, m.p. 224-226' dec. (lit. 225226'). Monoiodo-o-methyltyrosine.-Concentrated sulfuric acid (1 .0 cc.) and finely powdered iodine (1.48 9.) were added t o a suspension of o-methyltyrosine hydrogen sulfate' (3.40 g.) in alcohol (15 cc.). The reaction mixture was maintained a t 50" and treated slowly with 30% hydrogen peroxide solution (1.4 cc.) during ten minutes. The temperature was raised to 65' for five minutes, the solution then was diluted with water (10 cc.) and adjusted t o PH 7.5 with concentrated ammonia when crystallization of the product began. Ten per cent. aqueous sodium hydrogen sulfite (3.0 cc.) was added and the mixture was cooled. The white crystalline mass was collected, washed with alcohol, and air-dried (3.12 g., 84%, m.p. 220-221'). Recrystallized from water, the monoiodEo-methyltyrosine separated in colorless needles, m.p. 222 . Anal. Calcd. for Cl~H~zINOs.1/2H20: C, 36.3; H , 4.0; 1,38.5; N,4.2. Found: C,36.3; H,4.0; I,39.2; N,4.4. (6) All melting points are uncorrected. (7) L D . Behr and H. T. Clarke, THISJOURNAL, 64, 1630 (1932)
DEPARTMENT OF CHEMISTRY UNIVERSITY OF CALIFORNIA Los ANGELES,CALIFORNIA
The Chemistry of Nitroacetic Acid and its Esters. 111. The Synthesis of Tryptamine from Ethyl-a-
nitro-P-(3-indole)-propionate1 BY DOUGLAS A. LYTTLEA N D DAVIDI. WEISBLAT JUNE 27, 1955 RECEIVED
Ethyl a-nitro-~-(3-indole)-propionate (I) is a key intermediate in our synthesis of dl-tryptophan from ethyl nitroacetate2a or ethyl nitromalonate2b and (1) D. I. Weisblat a n d D . A. Lyttle, U. S. Patent 2,616,896. (2) (a) D. A. Lyttle and D. I . Weisblat, THISJOURNAL, 69, 2118 (1047); (b) 71, ,1079 (1949).
5748
NOTES
gamine. The scheme below represents a convenient method for the preparation of tryptamine from this compound (I)
Vol. 77
was prepared by recrystallizing the above material twice from ethanol; m . p . 242-248". Anal. Calcd. for ClnHwC1S2:C, fil.06; IT, 6.66; K, 14.25. Found: C, 61.33; H , 6.11; S,14.23. T h e reduction was also carried out under the same conditions in the presence of a slight excess of acetic acid. The solvent was removed under reduced pressure and the residual solid was recrystallized from ethanol to give 60% of pure tryptamine acetate, m.p. 134-135". Free base prcpared from the acetate melted at 114-11,j" ( i i i i ( ~ i r . ) . RESEARCH LABORATORIES THEUPJOHS COMPASY KALAMAZOO, MICHIGAX
/IJTCH~YHCOOC.H, 1, OH -
__f
Q\si
SO?
2, H +
I
H
I
I
H
I1
I
H
I11
Hydroxylation of Benzene in Aqueous Solution in Hydrolysis of I is carried out a t room temperature the Presence of Hydroxylamine Hydrochloride using alcoholic sodium hydroxide. A solid sodium BY JAS 0. KOSECNY salt is collected, dissolved in water and treated with RECEIVEDFEBRUARY 21, 1953 an excess of acid to give 3-(2-nitr0ethyl)-indolel~~ (11) in 83% yield. The melting points we found for The formation of o-nitrosophenol a t room tempertwo dimorphic forms of this compound (55.5-56.1' ature by the action of aqueous hydrogen peroxide, and 68.3-69.2') agree with those reported by Noland and Hartman3 (56.5-57' and 68-68.5'). Cat- hydroxylamine hydrochloride and copper sulfate on alytic reduction of I1 (Raney Ni) gave tryptamine, benzene or phenol has been described in the literature.l Other o-nitrosophenols have been prepared2 isolated as the hydrochloride, in 81.67, yield. in the same manner from numerous aromatic hyExperimental drocarbons, phenols and their derivatives. o-Xi3-(2-Nitroethyl)-indole (II).-A solution of sodium hy- trosophenols, formed in small quantities, are idendroxide (32.0 g., 0.8 mole) in 64 ml. of water was added to tified readily owing to their formation of charnc78.6 g. (0.30 mole) of ethyl a-nitro-P-(3-indole)-propionate teristic and intensely colored complexes' with the ( I ) in 200 ml. of ethanol. The resulting solution was allowed to stand a t room temperature for 44 hours. Solvent ions of transitional metals. The reaction has heen was removed under reduced pressure until the flask con- attributed to the formation of -SOH radicals and tltiiied a mass of solid, semi-crystalline material. Ethanol their attack on the organic molecule. I n order to (400ml.) was added, the mixture was slurried well and the explain the seemingly specific catalytic action of solid was collected on a filter. It was washed with ethanol, then with ether and dried on the filter. The solid was dis- copper ions, it has been postulated3 that they react solved in 600 ml. of water, the solution was cooled in ice and with and stabilize the free radicals. acidified by slowly adding 207, hydrochloric acid until the The present investigation revealed that ferrous pH was between 4 and 5 . Crystalline material began t o sulfate, cuprous chloride and metallic copper also separate and the flask was cooled at 4' overnight. T h e light pink crystals were collected, washed with water and catalyzed the reaction. Traces of o-nitrosophenol were found to be formed from benzene even in the dried under vacuum. The yield of 3-(2-nitroethyl)-indole mas 47.4 g. (83.27,). Material of analytical purity was ob- absence of a catalyst, and under such conditions tained from the "practical grade" product described above the reaction rate was increased by exposing the by dissolving i t in 200 ml. of ethanol, treating the solution with charcoal, filtering and adding 95 ml. of warm water t o solution to X-rays. Phenol, however, reacted the warm (60') filtrate. The flask was allowed to cool readily with the solution of hydroxylamine hydroslowly to room temperature and was then placed in the re- chloride and hydrogen peroxide. The formation of frigerator for 2 days. The large, sparkling plates were -NOH radicals in the system has so far remained collected on a filter and washed with two 40-1111. portions of 507, ethanol. T h e product, dried under vacuum, weighed conjectural. However, ample evidence is available concerning the formation of hydroxyl radicals 39.14 g. (68.5%) and melted at 55.5-56.1'. Two months later the melting point of the same material was 68.3-69.2'. from hydrogen peroxide by the action of short wave It was shown by analysis and infrared spectra t h a t these are r a d i a t i ~ n of , ~ copper5 and iron salts6 and metallic dimorphic forms. that silver and m e r ~ u r y . ~It has been shown4,G,8 Anal. Calcd. for Cl~Hl&~Op:C, 63.14; H, 5.30; N , such systems oxidize benzene to phenol. It also 14.73. Found: C, 62.80; H , 5.20; S,14.71. Tryptamine (3-(2-Aminoethyl)-indole)(III).-In a Parr has been shown9 that hydroxylamine hydrochloride rocking autoclave of 250-ml. void volume was placed 25.8 g. is oxidized to nitric acid by hydrogen peroxide, and of 3-(2-nitroethyl)-indole, 102 ml. of ethanol, approxi- it is very probable that nitrous acid is formed in the mately 3.8 g. of Raney nickel catalyst and hydrogen under reaction as an intermediate. p-Pc'itrosophenolYhas a pressure of 1520 p.s.i. at 23'. The autoclave was heated as rapidly as possible. Very little reduction occurred until been prepared by the action of these reagents on the temperature reached 85", when i t was complete in phenol, and the formation of o-nitrosophenol from about 3 minutes. Heating was discontinued, the tempera- phenol and nitrous acid is described in this work.
ture continued to rise to looo, and the autoclave was allowed to cool slowly to room temperature. Catalyst was removed by filtration and the filtrate was concentrated under vacuum to 75 ml. An excess of 10% alcoholic hydrogen chloride was added, causing crystalline material t o precipitate. The mixture was chilled a t 4' overnight, the crystals were collected on a filter and washed with a little cold ethanol, then ether. T h e light brown, crystalline material weighed 21.77 g. (81.6y0),m.p. 240" dec. An analytical sample (3) W. E. Noland and P. J. Hartman, THISJOURNAL, 76, 3227 (19Z4).
(1) 0. Baudisch, Science, 92, 336 (1940). (2) G. Cronheim, J. Oug. Cizem., 12, l ( 1 9 4 7 ) . (3) 0. Baudisch, Science, 108, 443 (1948). e, 869 (4) H. G. C. Bntes, h f . G. Evans and N. Cri, V n t z ~ ~ 166, (1950). ( 5 ) J, H. Baxendale, M. G. Evans a n d G. S . Park, T r a m . P a r a d a y Soc., 42, 155 (1946). (6) H. Loeb, G. Stein and D. Weiss, J . Chem. Soc., 2074 (15149). ( 7 ) J. Weiss, Trans. F a r a d a y .9or., 31, 1.547 ( 1 9 3 5 ) . (8) J. 0. Konecny, THISJ O U R N A L , 7 6 , 4U!4:3 (l!l.*14). (9) C . Wurster. B e r . , 20, 2631 (1887).
